Experience dependent plasticity of higher visual cortical areas in the mouse.

Experience dependent plasticity in the visual cortex is a key paradigm for the study of mechanisms underpinning learning and memory. Despite this, studies involving manipulating visual experience have largely been limited to the primary visual cortex, V1, across various species. Here we investigated the effects of monocular deprivation (MD) on the ocular dominance (OD) and orientation selectivity of neurons in four visual cortical areas in the mouse: the binocular zone of V1 (V1b), the putative "ventral stream" area LM and the putative "dorsal stream" areas AL and PM. We employed two-photon calcium imaging to record neuronal responses in young adult mice before MD, immediately after MD, and following binocular recovery. OD shifts following MD were greatest in LM and smallest in AL and PM; in LM and AL, these shifts were mediated primarily through a reduction of deprived-eye responses, in V1b and LM through an increase in response through the non-deprived eye. The OD index recovered to pre-MD levels within 2 weeks in V1 only. MD caused a reduction in orientation selectivity of deprived-eye responses in V1b and LM only. Our results suggest that changes in OD in higher visual areas are not uniformly inherited from V1.

Refinements to rodent head fixation and fluid/food control for neuroscience.

The use of head fixation in mice is increasingly common in research, its use having initially been restricted to the field of sensory neuroscience. Head restraint has often been combined with fluid control, rather than food restriction, to motivate behaviour, but this too is now in use for both restrained and non-restrained animals. Despite this, there is little guidance on how best to employ these techniques to optimise both scientific outcomes and animal welfare. This article summarises current practices and provides recommendations to improve animal wellbeing and data quality, based on a survey of the community, literature reviews, and the expert opinion and practical experience of an international working group convened by the UK's National Centre for the Replacement, Refinement and Reduction of Animals in Research (NC3Rs). Topics covered include head fixation surgery and post-operative care, habituation to restraint, and the use of fluid/food control to motivate performance. We also discuss some recent developments that may offer alternative ways to collect data from large numbers of behavioural trials without the need for restraint. The aim is to provide support for researchers at all levels, animal care staff, and ethics committees to refine procedures and practices in line with the refinement principle of the 3Rs.

Recruitment of frontal sensory circuits during visual discrimination.

A long-range circuit linking the medial frontal cortex to the primary visual cortex (V1) has been proposed to mediate visual selective attention in mice during visually guided behavior. Here, we use in vivo two-photon functional imaging to measure the endogenous activity of axons of A24b/M2 neurons from this region projecting to layer 1 of V1 (A24b/M2-V1axons) in mice either passively viewing stimuli or performing a go/no-go visually guided task. We observe that while A24b/M2-V1axons are recruited under these conditions, this is not linked to enhancement of neural or behavioral measures of sensory coding. Instead, A24b/M2-V1axon activity is associated with licking behavior, modulated by reward, and biased toward the sensory cortical hemisphere representing the stimulus currently being discriminated.

The separate and combined properties of the granular (area 29) and dysgranular (area 30) retrosplenial cortex.

Retrosplenial cortex contains two principal subdivisions, area 29 (granular) and area 30 (dysgranular). Their respective anatomical connections in the rat brain reveal that area 29 is the primary recipient of hippocampal and parahippocampal spatial and contextual information while area 30 is the primary interactor with current visual information. Lesion studies and measures of neuronal activity in rodents indicate that retrosplenial cortex helps to integrate space from different perspectives, e.g., egocentric and allocentric, providing landmark and heading cues for navigation and spatial learning. It provides a repository of scene information that, over time, becomes increasingly independent of the hippocampus. These processes, reflect the interactive actions between areas 29 and 30, along with their convergent influences on cortical and thalamic targets. Consequently, despite their differences, both areas 29 and 30 are necessary for an array of spatial and learning problems.

Cortical and Striatal Electroencephalograms and Apomorphine Effects in the FUS Mouse Model of Amyotrophic Lateral Sclerosis.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is characterized by degeneration of motor neurons resulting in muscle atrophy. In contrast to the lower motor neurons, the role of upper (cortical) neurons in ALS is yet unclear. Maturation of locomotor networks is supported by dopaminergic (DA) projections from substantia nigra to the spinal cord and striatum. OBJECTIVE: To examine the contribution of DA mediation in the striatum-cortex networks in ALS progression. METHODS: We studied electroencephalogram (EEG) from striatal putamen (Pt) and primary motor cortex (M1) in ΔFUS(1-359)-transgenic (Tg) mice, a model of ALS. EEG from M1 and Pt were recorded in freely moving young (2-month-old) and older (5-month-old) Tg and non-transgenic (nTg) mice. EEG spectra were analyzed for 30 min before and for 60 min after systemic injection of a DA mimetic, apomorphine (APO), and saline. RESULTS: In young Tg versus nTg mice, baseline EEG spectra in M1 were comparable, whereas in Pt, beta activity in Tg mice was enhanced. In older Tg versus nTg mice, beta dominated in EEG from both M1 and Pt, whereas theta and delta 2 activities were reduced. In younger Tg versus nTg mice, APO increased theta and decreased beta 2 predominantly in M1. In older mice, APO effects in these frequency bands were inversed and accompanied by enhanced delta 2 and attenuated alpha in Tg versus nTg mice. CONCLUSION: We suggest that revealed EEG modifications in ΔFUS(1-359)-transgenic mice are associated with early alterations in the striatum-cortex interrelations and DA transmission followed by adaptive intracerebral transformations.

Contrast adaptation and interocular transfer in cortical cells: A re-analysis & a two-stage gain-control model of binocular combination.

How do V1 cells respond to, adapt to, and combine signals from the two eyes? We tested a simple functional model that has monocular and binocular stages of divisive contrast gain control (CGC) that sit before, and after, binocular summation respectively. Interocular suppression (IOS) was another potential influence on contrast gain. Howarth, Vorobyov & Sengpiel (2009, Cerebral Cortex, 19, 1835-1843) studied contrast adaptation and interocular transfer in cat V1 cells. In our re-analysis we found that ocular dominance (OD) and contrast adaptation at a fixed test contrast were well described by a re-scaling of the unadapted orientation tuning curve - a simple change in response gain. We compared six variants of the basic model, and one model fitted the gain data notably better than the others did. When the dominant eye was tested, adaptation reduced cell response gain more when that eye was adapted than when the other eye was adapted. But when the non-dominant eye was tested, adapting either eye gave about the same reduction in overall gain, and there was an interaction between OD and adapting eye that was well described by the best-fitting model. Two key features of this model are that signals driving IOS arise 'early', before attenuation due to OD, while suppressive CGC signals are 'late' and so affected by OD. We show that late CGC confers a functional advantage: it yields partial compensation for OD, which should reduce ocular imbalance at the input to binocular summation, and improve the cell's sensitivity to variation in stereo disparity.

Opa1 Deficiency Leads to Diminished Mitochondrial Bioenergetics With Compensatory Increased Mitochondrial Motility.

Purpose: Retinal ganglion cells (RGCs) are susceptible to mitochondrial deficits and also the major cell type affected in patients with mutations in the OPA1 gene in autosomal dominant optic atrophy (ADOA). Here, we characterized mitochondria in RGCs in vitro from a heterozygous B6; C3-Opa1Q285STOP (Opa1+/-) mouse model to investigate mitochondrial changes underlying the pathology in ADOA. Methods: Mouse RGCs were purified from wild-type and Opa1+/- mouse retina by two-step immunopanning. The mitochondria in neurites of RGCs were labeled with MitoTracker Red for structure and motility measurement by time-lapse imaging. Mitochondrial bioenergetics were determined by the real-time measurement of oxygen consumption rate using a Seahorse XFe 96 Extracellular Flux Analyzer. Results: We observed a significant decrease in mitochondrial length in Opa1+/- RGCs with a remarkably higher proportion and density of motile mitochondria along the neurites. We also observed an increased transport velocity with a higher number of contacts between mitochondria in Opa1+/- RGC neurites. The oxygen consumption assays showed a severe impairment in basal respiration, Adenosine triphosphate-linked (ATP-linked) oxygen consumption, as well as reserve respiratory capacity, in RGCs from Opa1+/- mouse retina. Conclusions: Opa1 deficiency leads to significant fragmentation of mitochondrial morphology, activation of mitochondrial motility and impaired respiratory function in RGCs from the B6; C3-Opa1Q285STOP mouse model. This highlights the significant alterations in the intricate interplay between mitochondrial morphology, motility, and energy production in RGCs with Opa1 deficiency long before the onset of clinical symptoms of the pathology.

Stable Encoding of Visual Cues in the Mouse Retrosplenial Cortex.

The rodent retrosplenial cortex (RSC) functions as an integrative hub for sensory and motor signals, serving roles in both navigation and memory. While RSC is reciprocally connected with the sensory cortex, the form in which sensory information is represented in the RSC and how it interacts with motor feedback is unclear and likely to be critical to computations involved in navigation such as path integration. Here, we used 2-photon cellular imaging of neural activity of putative excitatory (CaMKII expressing) and inhibitory (parvalbumin expressing) neurons to measure visual and locomotion evoked activity in RSC and compare it to primary visual cortex (V1). We observed stimulus position and orientation tuning, and a retinotopic organization. Locomotion modulation of activity of single neurons, both in darkness and light, was more pronounced in RSC than V1, and while locomotion modulation was strongest in RSC parvalbumin-positive neurons, visual-locomotion integration was found to be more supralinear in CaMKII neurons. Longitudinal measurements showed that response properties were stably maintained over many weeks. These data provide evidence for stable representations of visual cues in RSC that are spatially selective. These may provide sensory data to contribute to the formation of memories of spatial information.

Spatial memory deficits initiated by agroclavine injection or olfactory bulbectomy in rats are characterized by different levels of long-term potentiation expression in the hippocampus.

Aim: To clarify whether long-term potentiation (LTP) is the mechanism underpinning mnemonic processes. Mathrials and methods: We studied LTP in hippocampal slices from rats whose spatial memory deficit was produced by either olfactory bulbectomy (OBX) or pretreatment with an ergot alkaloid, agroclavine. OBX is accompanied by cholinergic system inhibition whereas agroclavine predominantly activates dopaminergic mediation. The both have been shown to be involved in learning/memory and LTP mechanisms.Results: In OBX- vs. sham-operated rat, we have revealed significant reduction of LTP in hippocampal CA1 region. In contrast, no LTP differences in agroclavine- vs. vehicle-treated rats were observed. Conclusions: These results demonstrate that LTP expression in the hippocampus is dependent on the origin of spatial memory impairment. Furthermore, they suggest that pharmacological and neurodegenerative models of AD might be useful approach for discovery of both AD mechanisms and mixed pathology dementias.

Mammillothalamic Disconnection Alters Hippocampocortical Oscillatory Activity and Microstructure: Implications for Diencephalic Amnesia.

Diencephalic amnesia can be as debilitating as the more commonly known temporal lobe amnesia, yet the precise contribution of diencephalic structures to memory processes remains elusive. Across four cohorts of male rats, we used discrete lesions of the mammillothalamic tract to model aspects of diencephalic amnesia and assessed the impact of these lesions on multiple measures of activity and plasticity within the hippocampus and retrosplenial cortex. Lesions of the mammillothalamic tract had widespread indirect effects on hippocampocortical oscillatory activity within both theta and gamma bands. Both within-region oscillatory activity and cross-regional synchrony were altered. The network changes were state-dependent, displaying different profiles during locomotion and paradoxical sleep. Consistent with the associations between oscillatory activity and plasticity, complementary analyses using several convergent approaches revealed microstructural changes, which appeared to reflect a suppression of learning-induced plasticity in lesioned animals. Together, these combined findings suggest a mechanism by which damage to the medial diencephalon can impact upon learning and memory processes, highlighting an important role for the mammillary bodies in the coordination of hippocampocortical activity.SIGNIFICANCE STATEMENT Information flow within the Papez circuit is critical to memory. Damage to ascending mammillothalamic projections has consistently been linked to amnesia in humans and spatial memory deficits in animal models. Here we report on the changes in hippocampocortical oscillatory dynamics that result from chronic lesions of the mammillothalamic tract and demonstrate, for the first time, that the mammillary bodies, independently of the supramammillary region, contribute to frequency modulation of hippocampocortical theta oscillations. Consistent with the associations between oscillatory activity and plasticity, the lesions also result in a suppression of learning-induced plasticity. Together, these data support new functional models whereby mammillary bodies are important for coordinating hippocampocortical activity rather than simply being a relay of hippocampal information as previously assumed.

Loss of Midbrain Dopamine Neurons and Altered Apomorphine EEG Effects in the 5xFAD Mouse Model of Alzheimer's Disease.

Cognitive malfunction, synaptic dysfunction, and disconnections in neural networks are core deficits in Alzheimer's disease (AD). 5xFAD mice, a transgenic model of AD, are characterized by an enhanced level of amyloid-ß and abnormal neurotransmission. The dopaminergic (DA) system has been shown to be involved in amyloid-ß transformations and neuronal plasticity; however, its role in functional network changes in familial AD still remains unclear. In 5xFAD and non-transgenic freely moving mice, electroencephalograms (EEGs) were simultaneously recorded from the secondary motor cortex (MC), superficial layers of the hippocampal CA1 area (HPC), substantia nigra (SN), and ventral tegmental area (VTA). EEGs and their frequency spectra were analyzed before and after systemic injection of a DA receptor agonist, apomorphine (APO). In the baseline EEG from MC and HPC of 5xFAD mice, delta and alpha oscillations were enhanced and beta activity was attenuated, compared to control mice. In VTA and SN of 5xFAD mice, delta-theta activity was decreased and beta oscillations dominated. In control mice, APO suppressed delta activity in VTA to a higher extent than in MC, whereas in 5xFAD mice, this difference was eliminated due to attenuation of the delta suppression in VTA. APO increased beta activity in MC of mice from both groups while significant beta suppression was observed in VTA of 5xFAD mice. These mice were characterized by significant decrease of tyrosine hydroxylase immunopositive cells in both VTA and SN and of DA transporter in MC and hippocampal dentate gyrus. We suggest that the EEG modifications observed in 5xFAD mice are associated with alterations in dopaminergic transmission, resulting in adaptive changes in the cerebral networks in the course of familial AD development.

Plasticity in Adult Mouse Visual Cortex Following Optic Nerve Injury.

Optic nerve (ON) injury is an established model of axonal injury which results in retrograde degeneration and death of retinal ganglion cells as well anterograde loss of transmission and Wallerian degeneration of the injured axons. While the local impact of ON crush has been extensively documented we know comparatively little about the functional changes that occur in higher visual structures such as primary visual cortex (V1). We explored the extent of adult cortical plasticity using ON crush in aged mice. V1 function of the contralateral hemisphere was assessed longitudinally by intrinsic signal imaging and 2-photon calcium imaging before and after ON crush. Functional imaging demonstrated an immediate shift in V1 ocular dominance towards the ipsilateral, intact eye, due to the expected almost complete loss of responses to contralateral eye stimulation. Surprisingly, within 2 weeks we observed a delayed increase in ipsilateral eye responses. Additionally, spontaneous activity in V1 was reduced, similar to the lesion projection zone after retinal lesions. The observed changes in V1 activity indicate that severe ON injury in adulthood evokes cortical plasticity not only cross-modally but also within the visual cortex; this plasticity may be best compared with that seen after retinal lesions.

Midget retinal ganglion cell dendritic and mitochondrial degeneration is an early feature of human glaucoma.

Glaucoma is characterized by the progressive dysfunction and loss of retinal ganglion cells. However, the earliest degenerative events that occur in human glaucoma are relatively unknown. Work in animal models has demonstrated that retinal ganglion cell dendrites remodel and atrophy prior to the loss of the cell soma. Whether this occurs in human glaucoma has yet to be elucidated. Serial block face scanning electron microscopy is well established as a method to determine neuronal connectivity at high resolution but so far has only been performed in normal retina from animal models. To assess the structure-function relationship of early human glaucomatous neurodegeneration, regions of inner retina assessed to have none-to-moderate loss of retinal ganglion cell number were processed using serial block face scanning electron microscopy (n = 4 normal retinas, n = 4 glaucoma retinas). This allowed detailed 3D reconstruction of retinal ganglion cells and their intracellular components at a nanometre scale. In our datasets, retinal ganglion cell dendrites degenerate early in human glaucoma, with remodelling and redistribution of the mitochondria. We assessed the relationship between visual sensitivity and retinal ganglion cell density and discovered that this only partially conformed to predicted models of structure-function relationships, which may be affected by these early neurodegenerative changes. In this study, human glaucomatous retinal ganglion cells demonstrate compartmentalized degenerative changes as observed in animal models. Importantly, in these models, many of these changes have been demonstrated to be reversible, increasing the likelihood of translation to viable therapies for human glaucoma.

Digestion of the glycosaminoglycan extracellular matrix by chondroitinase ABC supports retinal ganglion cell dendritic preservation in a rodent model of experimental glaucoma.

Retinal ganglion cell dendritic atrophy is an early feature of glaucoma, and the recovery of retinal ganglion cell dendrites is a viable option for vision improvement in glaucoma. Retinal ganglion cell neurites are surrounded by a specialised glycosaminoglycan extracellular matrix which inhibits dendritic plasticity. Since digestion of the extracellular matrix by chondroitinase ABC has been reported to have neuro-regenerative and neuro-plastic effects within the central nervous system, we explored its potential for dendritic recovery in a rat model of ocular hypertension. Chondroitinase ABC was administrated intravitreally 1 week after ocular hypertension (a time point where dendritic atrophy has already occurred). Retinal ganglion cell dendritic morphology was unaffected by chondroitinase ABC in normal retina. In ocular hypertensive eyes retinal ganglion cells showed significantly decreased dendritic length and area under the Sholl curve with atrophy confined to higher order dendrites. These changes were not observed in chondroitinase ABC injected eyes despite similar total retinal ganglion cell loss (i.e. dendritic protection of surviving retinal ganglion cells). These data suggest that glycosaminoglycan digestion could have a therapeutic role in mitigating the effects of elevated pressure on retinal ganglion cell dendritic structure in glaucoma.

Spatial Memory Engram in the Mouse Retrosplenial Cortex.

Memory relies on lasting adaptations of neuronal properties elicited by stimulus-driven plastic changes [1]. The strengthening (and weakening) of synapses results in the establishment of functional ensembles. It is presumed that such ensembles (or engrams) are activated during memory acquisition and re-activated upon memory retrieval. The retrosplenial cortex (RSC) has emerged as a key brain area supporting memory [2], including episodic and topographical memory in humans [3-5], as well as spatial memory in rodents [6, 7]. Dysgranular RSC is densely connected with dorsal stream visual areas [8] and contains place-like and head-direction cells, making it a prime candidate for integrating navigational information [9]. While previous reports [6, 10] describe the recruitment of RSC ensembles during navigational tasks, such ensembles have never been tracked long enough to provide evidence of stable engrams and have not been related to the retention of long-term memory. Here, we used in vivo 2-photon imaging to analyze patterns of activity of over 6,000 neurons within dysgranular RSC. Eight mice were trained on a spatial memory task. Learning was accompanied by the gradual emergence of a context-specific pattern of neuronal activity over a 3-week period, which was re-instated upon retrieval more than 3 weeks later. The stability of this memory engram was predictive of the degree of forgetting; more stable engrams were associated with better performance. This provides direct evidence for the interdependence of spatial memory consolidation and RSC engram formation. Our results demonstrate the participation of RSC in spatial memory storage at the level of neuronal ensembles.

Animal models of amblyopia.

Unquestionably, the last six decades of research on various animal models have advanced our understanding of the mechanisms that underlie the many complex characteristics of amblyopia as well as provided promising new avenues for treatment. While animal models in general have served an important purpose, there nonetheless remain questions regarding the efficacy of particular models considering the differences across animal species, especially when the goal is to provide the foundations for human interventions. Our discussion of these issues culminated in three recommendations for future research to provide cohesion across animals models as well as a fourth recommendation for acceptance of a protocol for the minimum number of steps necessary for the translation of results obtained on particular animal models to human clinical trials. The three recommendations for future research arose from discussions of various issues including the specific results obtained from the use of different animal models, the degree of similarity to the human visual system, the ability to generate animal models of the different types of human amblyopia as well as the difficulty of scaling developmental timelines between different species.

Spatial summation across the visual field in strabismic and anisometropic amblyopia.

Ricco's area (the largest area of visual space in which stimulus area and intensity are inversely proportional at threshold) has previously been hypothesised to be a result of centre/surround antagonism in retinal ganglion cell receptive fields, but recent evidence suggests a sizeable cortical contribution. Here, Ricco's area was measured in amblyopia, a condition in which retinal receptive fields are normal, to better understand its physiological basis. Spatial summation functions were determined at 12 visual field locations in both eyes of 14 amblyopic adults and 15 normal-sighted controls. Ricco's area was significantly larger in amblyopic eyes than in fellow non-amblyopic eyes. Compared to the size of Ricco's area in control eyes, Ricco's area measured significantly larger in amblyopic eyes. Additionally, Ricco's area in the fellow, non-amblyopic eye of amblyopic participants measured significantly smaller than in control eyes. Compared to controls, Ricco's area was larger in amblyopic eyes and smaller in fellow non-amblyopic eyes. Amblyopia type, binocularity, and inter-ocular difference in visual acuity were significantly associated with inter-ocular differences in Ricco's area in amblyopes. The physiological basis for Ricco's area is unlikely to be confined to the retina, but more likely representative of spatial summation at multiple sites along the visual pathway.

Optophysiological Characterisation of Inner Retina Responses with High-Resolution Optical Coherence Tomography.

Low coherence laser interferometry has revolutionised quantitative biomedical imaging of optically transparent structures at cellular resolutions. We report the first optical recording of neuronal excitation at cellular resolution in the inner retina by quantifying optically recorded stimulus-evoked responses from the retinal ganglion cell layer and comparing them with an electrophysiological standard. We imaged anaesthetised paralysed tree shrews, gated image acquisition, and used numerical filters to eliminate noise arising from retinal movements during respiratory and cardiac cycles. We observed increases in contrast variability in the retinal ganglion cell layer and nerve fibre layer with flash stimuli and gratings. Regions of interest were subdivided into three-dimensional patches (up to 5-15 µm in diameter) based on response similarity. We hypothesise that these patches correspond to individual cells, or segments of blood vessels within the inner retina. We observed a close correlation between the patch optical responses and mean electrical activity of the visual neurons in afferent pathway. While our data suggest that optical imaging of retinal activity is possible with high resolution OCT, the technical challenges are not trivial.

Intrahippocampal Pathways Involved in Learning/Memory Mechanisms are Affected by Intracerebral Infusions of Amyloid-ß25-35 Peptide and Hydrated Fullerene C60 in Rats.

Primary memory impairments associated with increased level of amyloid-ß (Aß) in the brain have been shown to be linked, partially, with early pathological changes in the entorhinal cortex (EC) which spread on the whole limbic system. While the hippocampus is known to play a key role in learning and memory mechanisms, it is as yet unclear how its structures are involved in the EC pathology. In this study, changes in memory and neuronal morphology in male Wistar rats intrahippocampally injected with Aß25-35 were correlated on days 14 and 45 after the injection to reveal specific cognitive-structural associations. The main focus was on the dentate gyrus (DG) and hippocampal areas of CA1 and CA3 because of their involvement in afferent flows from EC to the hippocampus through tri-synaptic (EC → DG → CA3 → CA1) and/or mono-synaptic (EC → CA1) pathways. Evident memory impairments were observed at both time points after Aß25-35 injection. However, on day 14, populations of morphological intact neurons were decreased in CA3 and, drastically, in CA1, and the DG supramedial bundle was significantly damaged. On day 45, this bundle largely and CA1 neurons partially recovered, whereas CA3 neurons remained damaged. We suggest that Aß25-35 primarily affects the tri-synaptic pathway, destroying the granular cells in the DG supramedial area and neurons in CA3 and, through the Schaffer collaterals, in CA1. Intrahippocampal pretreatment with hydrated fullerene C60 allows the neurons and their connections to survive the amyloidosis, thus supporting the memory mechanisms.